Capillary ATPase activity has been thought to control water-electrolytes homeostasis in the brain. Endothelin-1 (ET-1) has recently been implicated in changes of blood-brain barrier (BBB) permeability. This report describes receptor (ET/A)-mediated stimulation of ouabain-sensitive (OS) and ouabain-insensitive (OI) 86Rb+ uptake (as a measure of Na+-K+-ATPase activity) by ET-1 and ET-3 in cultured rat brain capillary endothelial cells (BCEC). The uptake of 86Rb+ (0.2 micro Ci-well) was determined in confluent BCEC (grown in 96-microwell plates) incubated in M199 with HEPES for 5 min at room temperature. A concentration-dependent increase of 86Rb+ uptake induced by ET-1 or ET-3 (EC50=0.73 nM+/-0.17 and 12.89+/-3.69, respectively] was inhibited by the ET/A receptor antagonist (BQ-123) but not by the ET/B receptor antagonist (IRL 1038). Ouabain (ATPase inhibitor) and bumetanide (Na+-K+-Cl- cotransport inhibitor) decreased ET-1-stimulated 86Rb+ uptake into BCEC by 35% and 65%, respectively. Complete inhibition was seen with both of these agents. Similar results were observed with PMA, a PKC agonist. Ameloride [5-(N-ethyl-N-isopropyl], inhibitor of the Na+-H+ antiporter, decreased both the OS and OI ATPase induced by ET-1, suggesting a linkage of the Na+/H+ exchanger with Na+-K+- ATPase and Na+-K+-Cl- cotransport systems. The inhibition of ET-1-stimulated OS and OI 86Rb+ uptake with staurosporin (PKC antagonist) indicates that ET-1-induced ATPase activity is mediated by PKC. These results suggest that the effect of ET-1 on capillary OS and OI Na+-K+-Cl- systems may play a role in water-electrolyte disturbances in brain injury.